U.S. patent number 4,369,117 [Application Number 06/148,815] was granted by the patent office on 1983-01-18 for serum separating method and apparatus.
This patent grant is currently assigned to American Hospital Supply Corporation. Invention is credited to Fred K. White.
United States Patent |
4,369,117 |
White |
January 18, 1983 |
Serum separating method and apparatus
Abstract
A method and apparatus for separating serum from other blood
components in a centrifuge tube. The separator comprises a
cylindrical, laterally-expandable filter element composed of a
multiplicity of generally longitudinally-oriented fibers, 10 to 40
millimicrons in diameter, formed of a biologically inert polymeric
material having a specific gravity within the range of about 1.10
to 1.50 and bonded to adjacent fibers only at spaced-apart
junctures. The filter element has a diameter within the range of 9
to 15 millimeters, a length from 75 to 125 percent of its diameter,
and a bulk density of about 0.20 to 0.60 grams per cubic
centimeter. During centrifugation, the filter advances downwardly
from the mouth of the centrifuge tube, swelling laterally as serum
flows through the fine passageways running generally lengthwise of
the filter, and causing the descending filter to wipe against the
walls of the tube, pushing loose cells and fibrin ahead of it, and
finally compacting and restraining the fibrin-cell clot at the
bottom of the tube to permit subsequent decantation of the filtered
serum. In one embodiment, the filter is introduced into the tube by
means of a stretchable cap which serves as a carrier for the filter
prior to insertion and as a protective cover for the tube
thereafter.
Inventors: |
White; Fred K. (Miami, FL) |
Assignee: |
American Hospital Supply
Corporation (Evanston, IL)
|
Family
ID: |
22527514 |
Appl.
No.: |
06/148,815 |
Filed: |
May 12, 1980 |
Current U.S.
Class: |
210/782;
210/360.1; 210/508; 422/918 |
Current CPC
Class: |
B01D
35/00 (20130101); B01D 39/163 (20130101); G01N
33/491 (20130101); B01D 43/00 (20130101); B01L
3/5021 (20130101); B01D 39/18 (20130101) |
Current International
Class: |
B01D
39/16 (20060101); B01D 39/18 (20060101); B01D
43/00 (20060101); B01L 3/14 (20060101); B01D
35/00 (20060101); G01N 33/49 (20060101); B01D
021/26 () |
Field of
Search: |
;210/782,927,516,DIG.24,508 |
References Cited
[Referenced By]
U.S. Patent Documents
Primary Examiner: Adee; John
Attorney, Agent or Firm: Tilton, Fallon, Lungmus
Claims
I claim:
1. A filter element for insertion into a centrifuge tube containing
clotted whole blood for separating serum from the cellular and
fibrous components, comprising a laterally compressible and
expandable cylindrical body formed of a multiplicity of fibers
randomly oriented primarily in a longitudinal direction and
defining generally longitudinal flow passages therebetween; said
fibers being composed of an inert polymeric material of a specific
gravity between 1.10 to 1.50 and being bonded together at
spaced-apart junctures by a bonding agent; said fibers having
diameters within the range of about 10 to 40 micrometers and said
body having a bulk density of 0.20 to 0.60 grams per cubic
centimeter, a diameter of 9 to 15 millimeters, and a length of from
75 to 125% of said diameter; said polymeric material being
cellulose acetate and said binding agent being glycerol triacetate;
said glycerol triacetate being present in said filter element at a
percentage by weight no greater than 10%.
2. The filter element of claim 1 in which said glycerol triacetate
is present in said filter element at a percentage by weight no
greater than 4%.
3. The filter element of claim 1 in which said fibers have
diameters within the range of 15 to 25 micrometers.
4. The combination comprising an open-topped glass centrifuge tube
and a fibrous cylindrical filter element receivable in said tube;
said filter element being laterally compressible and expandable and
being formed entirely of a multiplicity of fibers randomly oriented
primarily in a longitudinal direction and bonded together only at
spaced-apart junctures to define a multiplicity of generally
longitudinal flow passages through said element; said fibers being
composed of an inert polymeric material of a specific gravity
between 1.10 to 1.50, and having diameters within the range of
about 10 to 40 micrometers; said element having a bulk density of
0.20 to 0.60 grams per cubic centimeter and a length of from 75 to
125% of the diameter thereof; said centrifuge tube having a
substantially uniform inside diameter throughout its length; said
diameter of said filter element in an unexpanded and uncompressed
state falling within 85 to 110% of said inside diameter of said
tube; said fibers of said filter element being bonded together at
spaced-apart junctures by a binding agent; said polymeric material
being cellulose acetate and said binding agent being glycerol
triacetate; said glycerol triacetate being present to an extent no
greater than 10% by weight of said filter element.
5. The combination of a serum filter element and a supporting cap
therefor, said filter element comprising a laterally compressible
and expandable cylindrical body consisting essentially of a
multiplicity of fibers randomly oriented primarily in a
longitudinal direction and bonded together only at spaced-apart
junctures to define generally longitudinal flow passages
therebetween; said fibers being composed of an inert polymeric
material of a specific gravity between 1.10 and 1.50, and having
diameters within the range of about 10 to 40 micrometers; said cap
being formed of resilient stretchable plastic material having a top
wall and an integral depending side wall extending about said
filter element; said side wall being provided with an
inwardly-projecting rib frictionally engaging said filter element
to retain the same within said cap when said side wall is
unstretched; said side wall being stretchable outwardly to an
extent sufficient to release said filter element; said side wall
including a downwardly and outwardly flared skirt portion
terminating in an enlarged opening at the bottom of said cap, said
opening being dimensioned to receive the upper end of a centrifuge
tube for enlarging said side wall and releasing said filter element
into such tube when said cap is forced downwardly over the tube's
upper end.
6. The combination of claim 5 in which said rib is annular and
defines an opening having a diameter smaller than the diameter of
said filter element in an uncompressed state.
7. The combination of claim 5 in which said filter element has a
bulk density within the range of about 0.20 to 0.60 grams per cubic
centimeter.
8. The combination of claim 5 in which said filter element when
undistorted has a diameter within the range of 9 to 15 millimeters
and a length of from said 75 to 125% of said diameter.
9. The combination of claim 5 in which said fibers of said filter
element are formed of a polymeric material having a specific
gravity within the range of 1.20 to 1.40.
10. The combination of a serum filter element and a supporting cap
therefor, said filter element comprising a laterally compressible
and expandable cylindrical body consisting essentially of a
multiplicity of fibers randomly oriented primarily in a
longitudinal direction and bonded together only at spaced-apart
junctures to define generally longitudinal flow passages
therebetween; said fibers being composed of an inert polymeric
material of a specific gravity between 1.10 and 1.50, and having
diameters within the range of about 10 to 40 micrometers; said cap
being formed of resilient stretchable plastic material having a top
wall and an integral depending side wall extending about said
filter element; said side wall being provided with an inwardly
projecting rib frictionally engaging said filter element to retain
the same within said cap when said side wall is unstretched; said
side wall being stretchable outwardly to an extent sufficient to
release said filter element; said side wall also including a
downwardly and outwardly flared skirt portion terminating in an
enlarged opening at the bottom of said cap; said opening being
dimensioned to receive the upper end of a centrifuge tube for
enlarging said side wall and releasing said filter element into
such tube when said cap is forced downwardly over the tube's upper
end; and a centrifuge tube having an inside diameter larger than
the diameter of said filter element and an outside diameter capable
of being received within said enlarged opening of said cap.
11. The combination of claim 10 wherein said centrifuge tube
includes an external lip at the upper end thereof; said side wall
of said cap being stretchable outwardly to an extent sufficient to
position said rib beneath said lip to latch said cap upon said
tube.
12. A method of treating a sample of blood to separate clear serum
therefrom, comprising the steps of collecting a sample of blood and
allowing the same to coagulate in a centrifuge tube to form a
fibrin-cell clot therein; then inserting a laterally-expandable,
porous and fibrous cylindrical filter element into said tube and
centrifuging said tube and its contents to cause said element to
descend towards the bottom of said tube behind said clot while
allowing serum to flow longitudinally upwardly through the pores of
said element; said element being composed of a multiplicity of
fibers randomly oriented primarily in a longitudinal direction and
formed of an inert polymeric material having a specific gravity
within the range of 1.10 to 1.50; said fibers having diameters
within the range of about 10 to 40 micrometers and said filter
element having a bulk density of about 0.20 to 0.60 grams per cubic
centimeter; and continuing centrifugation of said tube and its
contents until said filter element engages and compresses said clot
within the lower portion of said tube.
13. The method of claim 12 in which the specific gravity of said
polymeric material of said fibers falls within the range of 1.20 to
1.40.
14. The method of claim 12 in which said polymeric material is
selected from the group consisting of cellulose acetate, ethyl
cellulose, cellulose propionate, cellulose acetate butyrate, nylon,
and polyvinyl chloride.
15. The method of claim 14 in which adjacent fibers of said filter
element are bonded together at spaced-apart junctures.
16. The method of claim 15 in which said adjacent fibers are bonded
together at spaced-apart junctures by a binding agent selected from
the group consisting of glycerol triacetate, diethoxyethyl
phthalate, dimethoxyethyl phthalate, triethyl citrate, tributyl
citrate, tricresyl phosphate, glycerol tripropionate, triphenyl
phosphate, ethyl glycolate, acetyl triiso hexyl citrate, acetyl
triethyl citrate, dimethyl phthalate, diethyl phthalate, and
triethyl phosphate.
17. The method of claim 16 in which said polymeric material is
cellulose acetate and said binding agent is glycerol
triacetate.
18. The method of claim 12 in which said filter element expands
outwardly into wiping engagement with the inner surface of said
tube in response to the invasion of serum between the fibers
thereof upon commencement of centrifuging step.
19. A method of treating a coagulated sample of blood in an
open-topped centrifuge tube to separate clear serum from the clot
of fibrin and cellular components, comprising the steps of
inserting a laterally-expandable, porous, cylindrical filter
element into the open top of said tube and then centrifuging said
tube and its contents to cause said element to descend towards the
bottom of said tube behind said clot while allowing serum to flow
upwardly into and through said element; said element being formed
of a multiplicity of fibers randomly oriented primarily in a
longitudinal direction and bonded together only at spaced-apart
junctures; said fibers being composed of an inert polymeric
material having a specific gravity greater than any of the
constituents of said blood sample; said polymeric material of said
fibers being cellulose acetate and said fibers being bonded
together at said spaced-apart junctures by glycerol triacetate; and
continuing said centrifugation until said filter element engages
and compresses said clot within the lower portion of said tube.
20. A method of treating a sample of coagulated blood contained in
an open-topped centrifuge tube for the purpose of separating clear
serum from the fibrin-cellular clot therein, comprising the steps
of positioning a cap having a depending stretchable annular side
wall above the open top of said tube, said stretchable side wall
including a cylindrical upper portion, a downwardly flared skirt
portion, and an internal rib therebetween, said rib defining an
opening smaller than the inside of said tube when said side wall is
in an unstretched state, said rib engaging and releasably holding a
cylindrical filter element dimensioned to be axially received
within said tube through the open top thereof, then lowering said
cap over the open end of said tube to insert said filter element
into said tube and bring the inside surface of said skirt portion
and said rib into engagement with the outside of said tube at the
top thereof, then forcing said cap downwardly over said tube to
cause said side wall to stretch outwardly and release said filter
element within said tube.
21. The method of claim 20 in which said tube has a lip extending
about the open top thereof, and in which there is the further step
of continuing to force said cap downwardly over said tube until
said rib is located beneath said lip, thereby latching said cap
upon the upper end of said tube.
22. The method of claim 20 or 21 in which there is the further step
of centrifuging said tube and its contents to cause said filter
element to travel towards the bottom of said tube behind said clot,
while allowing serum to flow upwardly into and through said filter
element.
23. The method of claim 22 in which said filter element is formed
of a multiplicity of fibers randomly oriented primarily in a
longitudinal direction and bonded together at spaced-apart
junctures to define a multiplicity of longitudinal flow passages
between said fibers, said fibers being composed of an inert
polymeric material having a specific gravity greater than the
constituents of blood.
24. A method of treating a sample of coagulated blood contained in
an open-topped centrifuge tube for the purpose of separating clear
serum from the fibrin-cellular clot therein, comprising the steps
of positioning a cap having a depending stretchable annular side
wall above the open top of said tube, said stretchable side wall
including a cylindrical upper portion, a downwardly flared skirt
portion, and an internal rib therebetween, said rib defining an
opening smaller than the inside of said tube when said side wall is
in an unstretched state, said rib engaging and releasably holding a
cylindrical filter element dimensioned to be axially received
within said tube through the open top thereof, said filter element
being formed of a multiplicity of fibers randomly oriented
primarily in a longitudinal direction and bonded together at
spaced-apart junctures to define a multiplicity of longitudinal
flow passages between said fibers, said fibers being composed of an
inert polymeric material having a specific gravity within the range
of 1.10 to 1.50, said fibers having a diameter within the range of
10 to 40 micrometers, and said filter element having a bulk density
within the range of 0.20 to 0.60 grams per cubic centimeter, then
lowering said cap over the open end of said tube to insert said
filter element into said tube and bring the inside surface of said
skirt portion into engagement with the top of said tube, then
forcing said cap downwardly over said tube to cause said side wall
to stretch outwardly and release said filter element within said
tube, and centrifuging said tube and its contents to cause said
filter element to travel towards the bottom of said tube behind
said clot, while allowing serum to flow upwardly into and through
said filter element.
25. The method of claim 24 in which said filter element has a
length of from 75 to 125% of the diameter thereof.
Description
BACKGROUND
For a number of years, especially since the advent of automated
clinical analyses, there has been a need for simple and inexpensive
means to separate serum from the other constituents of coagulated
blood so that various diagnostic tests may be performed on the
serum without danger that particulates in the fluid may cause
malfunctioning and possible breakdown of the complex and costly
automated analysis equipment, or produce incorrect and misleading
test results, or both. Although the problems have been recognized
in the past, the proposed solutions to those problems have all had
major shortcomings.
In one type of centrifugally-activated system, a specially-prepared
centrifuge tube (which may also be stoppered, air-evacuated, and
serve as a blood collection tube) contains a mass of viscous
thixotropic gel at its lower (closed) end. The gel has a specific
gravity between that of serum (1.03) and the heavier cellular
components (1.09). Therefore, when a blood-filled tube is
centrifuged the contents will stratify with the gel assuming a
position between the serum (or plasma) and the cellular components.
See, for example U.S. Pat. No. 3,852,194. While such a system does
result in the formation of a barrier between serum (or plasma) and
the solid constituents, no filtering action, and no wiping or
cleaning of the tube surfaces above the equilibrium position of the
gel barrier, take place. Residual fibrin and cells may remain in
the serum layer, either clinging to the wall of the tube or
floating freely and, in either event, providing a source of
interference during subsequent testing of the serum (or plasma)
layer.
In an effort to reduce such problems, serum separators have been
devised in which the thixotropic gel is introduced from the
stoppered upper end of the tube, rather than from the closed lower
end, and migrates downwardly during centrifugation to assume its
equilibrium position at the serum-clot interface. (See U.S. Pat.
Nos. 3,986,962, 4,055,501, 3,957,654, 3,647,070, 4,012,325). Again,
however, in such a system the gel performs no significant wiping
and filtering actions. In general, such gel systems not only fail
to provide a clean separation between the liquid and solid
components of blood, but they also have the further disadvantages
of being relatively expensive and having only a limited
(approximately six months) shelf life.
In another type of centrifugally-activated serum separating device,
a piston is fitted into a centrifuge tube, the piston having a
specific gravity between that of serum (1.03) and red cells (1.09).
The piston has a resilient tube-contacting periphery and is also
provided with one or more openings so that during centrifugation it
will migrate from the upper end of the tube into an intermediate
equilibrium position at the serum-clot interface. See U.S. Pat.
Nos. 3,931,018, 3,951,801, 4,001,122. Such piston-type separators
are capable of performing effective separating and wiping actions
but only if the tubes are dimensioned to close tolerances, thereby
necessitating the use of special tubes which, added to the cost of
the separator, result in a relatively expensive assembly.
Other types of centrifugally-activated separators, some of which
combine gel barriers with piston-like structures, and others which
function as separators only when they are subjected to a further
treatment (such as heat treatment) after they have assumed their
equilibrium positions, are represented by U.S. Pat. Nos. 4,088,582,
3,909,419, 3,926,646, 3,920,557, and 3,919,085. Generally, most of
such centrifugally-activated separators, whether utilizing a gel,
or an apertured piston, or a combination of both, are automatically
interface-seeking because their specific gravity is set or adjusted
at about 1.06, at a point between the specific gravities of serum
(or plasma) and the heavier non-fluid blood components.
SUMMARY OF INVENTION
One important aspect of this invention lies in the discovery that a
superior interface-seeking separator may be obtained by utilizing a
fibrous laterally-expandable cylindrical filter element or plug
composed of an inert polymeric material having a specific gravity
significantly greater than all of the blood components, including
the heavier cellular components. Such a filter element, when
properly constructed and used in accordance with this invention,
provides effective wiping and filtering actions and produces a
clean separation between the serum layer and the particulate layer.
Furthermore, such a separator is relatively inexpensive and is
self-adjusting in use so that special tubes manufactured to closer
tolerances than standard tubes are not required.
A significant additional feature of this invention lies in
providing a method and apparatus which are operative to separate
the cellular and liquid components of blood only if such blood is
capable of coagulating under normal conditions. Thus, should a
sample to be tested have been drawn from a patient whose blood
resists coagulation because of hemophilia, anticoagulation therapy,
or any other reason, the serum separator of this invention, unlike
other separating devices and methods, will allow red cells to
remain in the upper stratum as a clear visual indication of the
patient's condition.
In a basic mode of practicing the invention, a sample of fresh
blood is first collected or placed in a tube capable of being
received by the tube carrier of a conventional hematological
centrifuge. Such tube may be the standard 100.times.13 or
100.times.16 (in millimeters) laboratory tubes commonly used for
blood collection. After an interval of approximately 30 minutes or
more to allow for coagulation, a laterally-expandable cylindrical
filter element is introduced into the mouth of the tube, and the
tube and its contents are then centrifuged to cuase the element to
migrate to the bottom of the tube behind the matrix or clot of
frbrin and cells. As the filter descends, serum flows upwardly
between the generally longitudinally oriented fibers causing the
loosely bonded fibers to separate slightly and increasing the
transverse dimensions of the filter as a whole. Consequently, the
cylindrical filter, which fit loosely into the mouth of the tube at
the outset, expands laterally and wipes the walls of the tube free
of loose cells and residual fibrin strands.
As the filter enters the lower portion of the tube it begins to
compress the clot because of the higher specific gravity of the
polymeric material from which the fibrous filter is formed.
Additional serum in the matrix of the clot is thereby extracted and
directed upwardly through the passageways of the filter. Downward
motion of the filter finally stops when it is resting on a tightly
packed cushion of cells and fibrin strands. Centrifugation is
discontinued and the clear serum is poured or otherwise removed
from the tube with the filter serving as a fixed barrier to prevent
intermixing of the supernatant with the trapped cells, fibrin
strands, and other particulates.
The filter is composed of a multiplicity of randomly-oriented
generally-longitudinal fibers formed of a substantially
biologically inert polymeric material having a specific gravity
within the range of about 1.10 to 1.50 (preferably 1.20 to 1.40)
and having fiber diameters within the range of about 10 to 40
(preferably 15 to 25) micrometers. The filter element should have a
bulk density of approximately 0.20 to 0.60 grams per cubic
centimeter and a diameter which permits it to be easily (rather
than tightly) received within the mouth of a straight-sided
cylindrical centrifuge tube. Such results may be achieved with a
filter which, in an uncompressed state, has a diameter falling
within the range of 85 to 110 percent of the tube's inside
diameter. The length of the cylindrical filter should be 75 to 125
percent of its diameter.
The filter element may be supplied to the user as part of an
assembly including a carrier cap. Such a cap is formed of flexible
and stretchable plastic material and has a side wall defining an
internal rib which engages and frictionally retains the filter
element. The side wall has an outwardly flared skirt portion
adapted to engage the mouth of a centrifuge tube and, upon such
engagement, to help pilot the filter into the tube and, as
insertion progresses, to expand outwardly causing the rib of the
cap to release its hold on the filter. Insertion of the filter
element is completed when the deformed cap encloses the upper
portion of the tube and the internal rib assumes a latching
position beneath the bead or lip of the centrifuge tube. The
stretchable cap therefore serves as a filter retainer during
transit, storage, and handling, a shield which performs a
protective and guiding function as the filter element is inserted
into the mouth of a centrifuge tube, and a cover which is capable
of automatically releasing the filter element and then latching
onto and sealing the mouth of the tube for subsequent processing of
the blood sample.
Other objects, features, and advantages of the invention will
become apparent from the specification and drawings.
DRAWINGS
FIG. 1 is a perspective view of a filter and cap assembly embodying
the invention.
FIG. 2 is an exploded perspective view showing the filter, cap, and
a portion of a conventional centrifuge tube.
FIG. 3 is a greatly enlarged cross sectional view of a portion of
the fibrous filter.
FIG. 4 is an enlarged longitudinal sectional view of the filter-cap
assembly.
FIG. 5 is a sectional view similar to FIG. 4 but illustrating the
relationship of the assembly with a centrifuge tube as the filter
is being inserted into the tube.
FIG. 6 is similar to FIG. 5 but shows the outward expansion of the
skirt of the cap as filter insertion progresses.
FIG. 7 is a longitudinal sectional view similar to FIGS. 4-6 but
showing the filter in inserted position and the deformed cap in
latched condition.
FIGS. 8 and 9 somewhat schematically depict the action of the
filter during centrifugation.
FIG. 10 illustrates the function of the filter as a
particulate-retaining barrier during pour-off of the serum from the
tube.
DETAILED DESCRIPTION
Referring to FIGS. 1-3, the numeral 10 generally designates a
fibrous filter element for use in the centrifugally-activated
separation of serum from other components of a blood sample. Cap 11
may be used as a carrier for the filter, to aid in inserting the
filter into a blood-collection centrifuge tube 12, and to seal the
open top of the tube when insertion of the filter element is
completed.
Centrifuge tube 12 is a conventional open-topped glass tube of the
type commonly used for the collecting and centrifuging of blood
samples. Such a tube is presently commercially available in two
sizes, specifically, 13.times.100 and 16.times.100, the approximate
outside dimensions measured in millimeters. Ordinarily, such tubes
are supplied to users in evacuated condition with piercable
stoppers (not shown) sealing their open ends. Because the tube is
air evacuated, the drawing of a blood sample from a patient
(utilizing a syringe equipped with a double-ended needle, one end
which is inserted into a vein and the other which is then driven
through the piercable stopper) is facilitated. Such a procedure for
collecting a blood sample, and the elements used for that purpose,
are well known and widely used, and need not be described in
further detail herein. It is to be noted, however, that while tube
12 would normally function as a blood collection tube as well as a
centrifuge tube, only the latter function is considered
particularly relevant with respect to the present invention since,
quite conceivably, the blood sample might be collected by means of
a standard plunger-equipped syringe and then transferred to tube
12.
In any event, tube 12 is cylindrical except for its rounded bottom
end 12a (FIG. 10) and a slight but definite lip or enlargement 12b
extending about the tube's mouth. The interior surfaces of the tube
must be clean and free of lubricants and other coatings.
The filter element or plug 10 takes the form of a laterally
compressible and expandable porous cylindrical body formed of a
multiplicity of fibers 13 randomly oriented primarily in a
longitudinal direction and defining a multiplicity of longitudinal
flow passages 14 therebetween. The fibers 13 are composed of an
inert polymeric material having a specific gravity within the range
of about 1.10 to 1.50, the preferred range being 1.20 to 1.40. The
term "insert" is used herein to mean a stable material that will
not react with blood or with reagents and devices employed in the
collection, storage, and analysis of blood, in any way that might
alter the results of any of the variety of known tests for organic
and inorganic constituents of serum. While cellulose acetate has
been found particularly effective, other polymeric materials may
also be used as, for example, ethyl cellulose, cellulose
propionate, cellulose acetate butyrate, nylon (polyamides),
polyvinyl chloride, and various copolymers of such materials, all
formulated to have a specific gravity within the range of 1.10 to
1.50.
The fibers of the filter element have diameters within the range of
about 10 to 40 micrometers (.mu.m), the preferred range being 15 to
25 .mu.m, and should be randomly oriented primarily in a
longitudinal direction. The phrase "randomly oriented in a
longitudinal direction" is used herein to have the same meaning as
disclosed in the prior art (see, for example, U.S. Pat. No.
3,111,702); that is, as a description of a condition of a body of
fibers which are as a whole longitudinally aligned and generally
parallel, but which nevertheless do have portions which are not in
precise parallel alignment with adjacent fibers and which therefore
contact such adjacent fibers at spaced-apart junctures or points of
contact. Any given fiber in the cylindrical filter element 10
extends from one end face of the element to the opposite end face
thereof; however, in doing so such fiber is not necessarily
straight or parallel with adjacent fibers.
The fibers are held weakly or loosely together by a suitable binder
which, like the fibers themselves, must be of a composition and
extent so as not to interfere with the operation of the filter and
with the test results. The binder simply functions to hold adjacent
fibers together at their spaced-apart junctures or points of
contact. The filter element must remain readily compressible and
expandable, and the multiplicity of the fine passageways extending
longitudinally through the element must not be occluded by the
binder.
Where cellulose acetate is used as the fiber material, glycerol
triacetate has been found to be particularly effective as the
binder material. Glycerol triacetate is commonly used as a binder
in the manufacture of other products, is considered ingestible and
harmless, and has a specific gravity within the range of 1.15 to
1.16. If present in bound form (i.e., not as free or unattached
particles within the fibrous structure) in an amount not exceeding
10% by weight of the filter element, and preferably at a level not
exceeding 4%, it has been found that glycerol triacetate, when used
as the binding agent for securing the fibers together at the nodes
where adjacent fibers intersect, does not interfere with
conventional tests for serum constituents such as calcium,
inorganic phosphorus, glucose, blood urea nitrogen, uric acid,
cholesterol, alkaline phosphatase, lactic dehydrogenase, albumin,
and the like. Glycerol triacetate may also be used when materials
other than cellulose acetate are used for the fibers and,
conversely, other suitable binders may be used to bond the fibers
(of any selected composition) together. Other binders include
diethoxyethyl phthalate, dimethoxyethyl phthalate, triethyl
citrate, tributyl citrate, tricresyl phosphate, glycerol
tripropionate, triphenyl phosphate, ethyl glycolate, acetyl triiso
hexyl citrate, acetyl triethyl citrate, dimethyl phthalate, diethyl
phthalate, and triethyl phosphate.
Processes that may be used in the manufacture of filamentary tow
from which filter elements of this invention may be made are
already known in the art. Reference may be had to U.S. Pat. Nos.
3,095,343 and 3,111,702. The filter elements, whether manufactured
by such procedure or by any other suitable technique, should have a
diameter within the range of 9 to 15 millimeters, a length of from
75 to 125% of their diameter, and a bulk density within the range
of 0.20 to 0.60 grams per cubic centimeter.
Within the range given, the particular diameter selected for the
filter elements depends on the size of the tubes with which they
are to be used. The filter elements should be small enough in
diameter to be easily insertable into the tubes without first
radially compressing such filters, yet must be large enough so that
in use, during centrifugation, the filter elements will expand
outwardly to produce an effective wiping action against the inside
surfaces of the tubes. A tight fit at the time of insertion should
be avoided because such frictional engagement, increased by the
expansive forces exerted during centrifugation, may prevent
downward movement of the filter elements towards the lower ends of
the tubes during centrifugation. The desired relationship between a
filter element and a tube at the time of insertion may be
characterized as a loose or free sliding fit, it being understood
that such a relationship may be achieved, despite substantial
manufacturing tolerance ranges for both the tubes and the elements,
because of the compressibility of such elements. For example,
standard centrifuge tubes of 13.times.100 outside dimensions (in
millimeters) have been found to have a considerable range of inside
diameters averaging approximately 10.5 millimeters and, for use
with such tubes, it has been found that filter elements averaging
approximately 9.9 to 10.0 millimeters, with a variance either way
of 0.4 millimeters, produce particularly effective results. In
general, the diameter of the filter in an uncompressed state should
fall within the range of 9 to 15 millimeters and should be 85 to
110 percent of the inside diameter of the centrifuge tube.
Filter length is important to insure adequate filtering and wiping
actions and to prevent the filters from becoming tipped and canted
within the tubes. Filters longer than approximately 125% of their
diameter retain excessive volumes of serum, whereas filters shorter
than approximately 75% of their diameter tend to be unstable within
the centrifuge tubes, are likely to wipe inadequately or unevenly,
and tend to allow fibrin strands and particulates to pass
therethrough into the serum layer.
Cap 11 is fabricated from a soft, readily deformable and
stretchable plastic such as low density polyethylene, silicone
rubber, polyvinyl chloride, or other polymeric materials having
their similar properties. The cap has an imperforate end wall 15
and a side wall 16, the latter including a generally cylindrical
upper portion 17 and an outwardly and downwardly lower portion or
skirt 18. As shown in FIG. 4, the upper cylindrical portion 17 is
relatively thin. Not only is the skirt 18 of thicker cross section,
but the dimensional difference appears as an annular internal rib
or shoulder 19 at the juncture of the two portions. The internal
diameter of the rib 19, that is, the diameter of the opening
defined by that rib when the cap is in an untensioned or
unstretched state, is smaller than the diameter of filter element
10 and the inside diameter of tube 12. The filter element is
therefore frictionally retained in the cap as depicted in FIG. 4,
with rib 19 causing slight inwardly deformation of the filter
10.
The flared skirt is dimensioned so that the inside diameter at its
open end approximates the outside diameter of centrifuge tube 12. A
user, gripping the cap-filter assembly by the cylindrical outer
surface of the cap may easily direct the lower end of the filter
into the mouth of the tube, bringing the lower surfaces of the
skirt 18 into engagement with the lip of the tube (FIGS. 5 and 6).
An annular flange 18a projecting outwardly from the lower end of
the skirt may be provided to assist a user in orienting the cap in
relation to the tube and to help protect against direct finger
contact with the mouth of the tube, and with the sample of blood
contained within the tube, as the cap-filter assembly is lowered.
As force is exerted to continue the downward movement of the cap
and filter, the rounded lip 12b of the tube bears against the
sloping inside surface of the skirt 18 and causes outward expansion
of the skirt (FIG. 6). At the same time, the filter element 10
proceeds further into the mouth of the tube and, because of the
outward expansion or deformation of the side wall of the cap, rib
19 releases its hold on the filter element. As the cap is forced
even further downwardly, the rib-providing portion of the cap
expands outwardly to accommodate the upper end of tube 12 until rib
19 finally clears lip 12a and snaps into latching position beneath
that lip (FIG. 7). It has been found that the tension of the cap,
as the rib 19 snaps into its latching position, produces an audible
sound to signal to the user that the filter element is fully
inserted and a sealing of the tube's open end has been
effected.
While cap 11 is valuable as a means for handling filter element 10,
introducing that element into a centrifuge tube 12, and sealing
that tube while simultaneously releasing the filter element within
the tube, it is not essential that the cap be used as part of the
operation of the filter element. If a user is willing to accept
increased risks of exposure and contamination, and the further
possibility that direct finger contact with a filter might have
some effect on test results, he might simply insert such filter
elements into the centrifuge tubes using his fingers to make direct
contact with such elements. On the same basis, subsequent handling
of the filter-equipped tube, and centrifuging of that tube and its
contents, may be performed without any seal or cap at the tube's
upper end. Regardless of whether cap 11 is or is not utilized,
filter element 10 will be introduced into the open end of tube 12
only after a fresh sample of blood has already been received in
that tube and only after coagulation of that blood has already
occurred. Where tube 12 also functions as a blood collection tube,
it has been found desirable to leave the original stopper in place
following collection until after full coagulation has taken place.
An interval of at least 30 minutes is generally required.
Thereafter, the original stopper is removed, filter element 10 is
inserted into the mouth of the tube, and the tube and its contents
are centrifuged at rotational speeds commonly used in clinical
laboratories for the centrifugation of blood. Quite typically,
centrifugation would be performed at a relative centrifugal force
(rcf) of 1100 or more for an interval of approximately 10
minutes.
As centrifugation commences, the cellular components of the blood
are dispersed in the serum and are loosely entrapped in a three
dimensional mesh of fibrin strands. This matrix or clot is then
relatively free to descend, such descent being restrained primarily
by weak bonds between the fibrin and the glass surfaces of the tube
and by the elasticity of the fibrin which tends to resist
deformation. As the clot matrix begins to descend, serum is
displaced upwardly through the voids between the cells and fibrin.
The clot concentrates at the lower end of the tube where, in the
absence of the filter element as part of the system, such clot
would retain a substantial volume of serum trapped in the voids
between the cells and fibrin. During its descent the clot would,
again in the absence of filter element 10, leave behind free fibrin
strands, cellular materials, and other particulates in the serum,
and other fibrin strands clinging to the inside surfaces of the
tube.
Descent and partial compaction of the clot takes place in advance
of descent of the filter element 10. The filter element moves
relatively slowly into the serum at the serum-air interface, the
serum displacing air from the porous filter and causing outward
expansion of the filter element into sliding frictional engagement
with the inside surface of the tube. As serum displaces air from
the filter element, the apparent density of the filter increases.
The filter element descends, wiping the walls of the tube and
pushing ahead loose cells and detached fibrin. The rate of descent
of the filter element is believed to be limited largely by friction
between the element and the walls of the tube and by fluid friction
between the serum and the surfaces defining the minute passageways
through the body of the element. The relationship between the
descending clot and the trailing filter element during the early
phase of centrifugation is somewhat schematically depicted in FIG.
8, where 20a represents the clarified serum, 20b designates the
fibrin-cell matrix, arrows 21 and 22 indicate the direction of
movement of the fibrin-cell matrix and the filter element,
respectively, arrows 23 represent the direction of flow of serum
through the filter element, and arrow 24 indicates the lateral
expansion of the filter element, and the forceful wiping contact
between that element and the inside surface of tube 12, occasioned
by resistance to the flow of serum through the element's minute
longitudinal passages.
Ultimately, the filter element contacts the clot or fibrin-cell
matrix 20b, such contact occurring well after the filter element
enters the lower half of the tube. Since the specific gravity of
the material from which the filter is formed is substantially
greater than that of the blood's cellular components, the filter
element begins to compress the fibrin-cell matrix 20b to squeeze
additional amounts of serum therefrom. The fact is supported by the
observation that the yield of serum as a result of centrifugation
is greater when a filter element is used, in comparison with a
straight-spin down without use of a filter element, even where the
filter element is formed of a hydrophilic material (such as
cellulose acetate) and therefore tends to retain a volume of serum
in its interstices. Downward motion of the filter element 10
eventually stops with the element resting on a tightly packed
cushion of fibrin and cells (FIG. 10). Following removal of cap 11
from the tube, the supernatant serum 20a may be poured or pipetted
from the tube, the filter element 10 remaining in place as a
barrier to prevent re-mixing of the serum and cellular
components.
While the filter element does perform a filtering function as well
as a wiping action, calculations reveal that the pores of
passageways extending generally longitudinally through the element
are large enough to permit red cells, which are highly deformable,
to pass therethrough under certain conditions without rupture of
the cell membranes. The fact that such cells do not do so under
normal operating conditions is believed to be attributed in a large
part to the lag in the descent of the filter element 10 with
respect to clot 20b, the relatively slow descent of the filter
element occasioned by the frictional forces generated between
element 10 and the inside surface of the tube, the lateral
expandability of the element resulting from the longitudinal
orientation of the fibers and the limited bonding therebetween, and
the length of the filter element in relation to its diameter. It is
to be noted that if a blood sample fails to clot prior to
centrifugation as, for example, where the sample has been drawn
from a hemophiliac or from a patient on anticoagulant therapy, red
cells do pass into and through filter element 10 and their presence
in the fluid above the filter element following centrifugation, and
the bottoming of the filter element in the tube (with no visible
red cells therebelow), clearly signal the special circumstance
presented.
The following example of a preferred embodiment and mode of
practicing the invention is given for purposes of illustration.
EXAMPLE
The performance of filter elements embodying this invention, and
the effectiveness of the method of using such filter elements, were
evaluated for 32 standard blood chemistry tests. The analytical
equipment used for conducting such tests included a SMA 12/60
analyzer (Technicon Instruments Corporation, Tarrytown, N.Y.) for
calcium, inorganic phosphorus, glucose, blood urea nitrogen, uric
acid, cholesterol, total protein, albumin, total bilirubin,
alkaline phosphatase, lactic dehydrogenase, and serum
glutamate-oxaloacetate transaminase, a duPont ACA (duPont,
Wilmington, Del.), for assaying values for carbon dioxide,
creatinine, creatine phosphokinase, chloride, lipase, amalase,
pseudocholinesterase, magnesium, iron, acid phosphatase,
triglyceride, glutamate-oxaloacetate transaminase and lactic acid,
and an Auto Analyzer II (Technicon Instruments Corporation,
Tarrytown, N.Y.) for sodium, potassium, and phosphorus. Four tubes
of blood were collected from each of eight donors using
16.times.100 millimeter evacuated collection tubes in accordance
with the manufacturer's instructions (Becton-Dickinson, Rutherford,
N.J.). One specimen from each donor was processed using a filter
element embodying the invention, as described below, a second
specimen was processed using a commercial gel separator (Sure-Sep
II Separator, General Diagnostics, Morris Plains, N.J.), a third by
using a commercial rubber disk separator (Glasrock Filter Sampler,
Glasrock Products, Inc., Fairburn, Ga.), and the fourth by using no
separator at all.
The filter elements embodying the invention were composed of
generally longitudinally-oriented cellulose acetate fibers bonded
together at spaced-apart junctures by a glycerol triacetate binder.
Prior to use the filter elements were heated to a temperature of
approximately 140.degree. C. to volatilize and remove free unbonded
glycerol triacetate therefrom. The residual bound glycerol
triacetate securing the fibers together at their crossing nodes was
determined to be less than 8% by weight.
Each filter was cylindrical in configuration with a diameter of
approximately 13.6 millimeters, a length of approximately 11.0
millimeters, and a bulk density of 0.232 grams per cubic
centimeter. The fibers were estimated to have an average diameter
of approximately 0.25 micrometers and a specific gravity of 1.32.
The average inside diameters of the blood-collecting centrifuge
tubes was determined to be about 13.6 millimeters.
The compressible fibrous filter elements were releasably supported
by stretchable filter caps similar to the caps depicted in FIGS. 1
and 4, such caps being formed from low density polyethylene (NA270,
USI, New York, N.Y.). The inside diameter of such caps, measured at
the internal ribs thereof when such caps were unstretched, was
approximately 13.2 millimeters.
The procedure for those specimens to be processed using the
cellulose acetate filters consisted of allowing each tube with a
freshly-drawn specimen to stand approximately 30 minutes for
clotting, then removing the rubber stopper from the blood
collection tube and inserting a filter into the open end of the
tube utilizing the filter cap as the element-inserting device, then
forcing the cap downwardly over the end of the tube to secure the
stretched cap in place with the lower end of the filter element
spaced just above the surface of the sample, then centrifuging the
tube and its contents at 1100 rcf for 10 minutes, then inspecting
the tube and its contents for hemolysis, barrier condition and
serum appearance, and finally pipetting amounts of serum from the
tube for the various blood chemistry tests. The procedures for
those specimens to be processed with commercial gel separators and
rubber disk separators were in accordance with the manufacturer's
instructions. All samples were compared for serum yield.
From visual inspections of the tubes containing the cellulose
acetate filters, following centrifugation of those tubes, it was
found that the fibrous filters caused no apparent hemolysis. In
each instance, the fibrous filter was found to be resting upon a
compacted fibrin-cell maxtrix at the lower end of the tube, having
wiped the inside surfaces of the tube during its descent to remove
fribrin and other particulate matter therefrom. The serum layer in
each tube in which a fibrous filter was used was clear and
substantially fibrin-free. In some cases, fibrin would penetrate
through the filter into the serum; however, the serum could still
be decanted because the fibrin would remain firmly attached to the
filter. Serum yield measurements showed that those specimens
processed with the fibrous filters provided a yield that was
essentially the same as those specimens processed with gel
separators, and an average of 15% greater than those samples
processed with rubber disk filters. In all instances in which the
fibrous filters were used, serum could be easily poured from the
tubes without dislodging the filter barriers from those tubes.
The data from the blood chemistry tests was analyzed statistically
using the paired t-test method where the bias, standard deviation,
t-value, and P-value were calculated. Most of the tests showed no
statistically significant differences (biases). In those tests
which showed statistically significant differences, such
differences were judged to be small and not clinically significant.
In such latter group were alkaline phosphatase, carbon dioxide,
cholesterol, SPK, glucose, LDH, sodium, total protein, uric acid,
and potassium. In total, no clinically significant differences were
observed between the specimens processed with the fibrous cellulose
acetate filters and those processed with no separators at all. Use
of the fibrous filters did appear to give significantly better
results in many analytical values than the use of gel or rubber
disk separators.
While in the foregoing I have disclosed an embodiment of the
invention in considerable detail for purposes of illustration, it
will be understood by those skilled in the art that many of these
details may be varied without departing from the spirit and scope
of the invention.
* * * * *